Methods of treating biliary lithiasis

ABSTRACT

The methods of this invention are useful for the treatment of biliary lithiasis in mammals. The methods involve administering to mammals certain fluorinated derivatives of esters of phenoxy isobutyric acid. While mammals subjected to a lithogenous regime developed cohesive doubly refractive crystals, mammals on the same regime and treated by the methods of this invention developed no lithiasis.

Q United States Patent 1 [111 3,860,721

Bulteau Jan. 14, 1975 [5 METHODS OF TREATING BILIARY [58] Field of Search 424/308. 3] l LITHIASIS [75] Inventor: Grard C. Bulteau, Paris, France References Cited [73] Assignee: Societe DEtudes Scientifiques et UNITED STATES PATENTS Industrielles de LIle-De-France, 3,549,691 12/1970 Leigh et a1 424/308 X Paris, France Examiner-Frederick Attorney, Agent, or firm-Nolan & Baxley [21] Appl. N0.: 359,678

Related US. Application Data [57] ABSTRACT Division of N 1 y 11, 1972, The methods of this invention are useful for the treatgli f, fg 7 l b g g g ment of biliary lithiasis in mammals. The methods inarc one ommuanon'm'part O volve administering to mammals certain fluorinated Sept 1968 derivatives of esters of phenoxy isobutyric acid. While mammals subjected to a lithogenous regime developed [30] Forelgn Apphcatlon Pnonty Data cohesive doubly refractive crystals, mammals on the NOV. 22, France same regime and the of invention developed no lithiasis. [52] US. Cl. 424/308 51] rm. Cl A6lk 27/00 6 clalm N0 Drawings METHODS OF TREATING BILIARY LITHIASIS This application is a division of U.S. patent application Ser. No. 270,626, filed July 11, 1972, now U.S. Pat. No. 3,790,681. The application Ser. No. 270,626 is a continuation-in-part of U.S. patent application, Ser. No. 124,957, tiled Mar. 16, 1971, now abandoned. The U.S. patent application Ser. No. 124,957 is a continuation'in-part of U.S. patent application Ser. No. 760,973 filed Sept. 19, 1968, now U.S. Pat. No. 3,632,629.

This invention relates to methods of treating biliary lithiasis in mammals. The methods comprise administering to mammals, orally or parenterally such as subcutaneously or intramuscularly, fluorinated derivatives of the esters of phenoxy isobutyric acid. The daily dosage is within the range of to 360 mg/kg of body weight of the mammal treated, and preferably for man within the range of 140 to 280 mg/kg of body weight, depending upon the severity of the case.

The fluorinated derivatives of the esters of phenoxy isobutyric acid employed in the methods of this invention have the formula:

in which R is an alkyl radical of 1 to 5 carbon atoms either straight or branched chain; X is trifluoromethyl, trifluoromethylthio, trifluoromethylsulfinyl or trifluoromethylsulfonyl.

The process of preparation of the fluorinated derivatives of the esters of the phenoxy isobutyric acid employed in the methods of this invention consists in starting with a substituted phenol in combination with a'radical such as trifluoromethyl, trifluoromethylthio, trifluoromethylsulfinyl, trifluoromethylsulfonyl; treating it with chloroform and acetone in the presence of potash so as to obtain the corresponding phenoxy isobutyric acid. This is then esterifled by an alcohol of low molecular weight such as methanol, ethanol, propanol, isopropanol, etc. in the presence of a catalyzing agent. The ethyl esters are particularly effective because of their high cholesterol control properties.

Typical examples of the derivatives of esters of phenoxy isobutyric acid utilized in the methods of this invention are as follows:

EXAMPLE I ETHYL-P-TRIFLUOROMETHYLPHENOXYISOBU- TYRATE dium hypobromide for 48 hours. When the reaction is completed, the product is dried, washed with water and recovered with methanol. 112 G of pnitrotribromotoluene are obtained. (Yie1d230percent m.p. 86C) Stage B P-Trifluoromethylnitrobenzene Into a 1 liter flask equipped with a small distillation column joined to a descendant refrigerant, 112 g (0.3 mole) of p-nitrotribromotoluene and g of pulverized antimony trifluoride are introduced. The flask is heated. When the highly exothermic reaction is completed, the product is distilled and is then extracted with ethyl ether. 40 G of p-trifluoromethylnitrobenzene are obtained. (Yield: 70 percent m.p. 40C) Stage C P-Trifluoromethylaniline Into a 1 liter three-necked flask equipped with a mechanical agitator, a thermometer and a dropping funnel are introduced 40 g of p-trifluoromethylnitrobenzene (0.2 mole) and 120 ml of absolute ethanol. There is then poured in under agitation a solution of 240 g of stannous chloride in 200 ml of hydrochloric acid (d: 1.19). When the reduction is completed, the mixture is neutralized with soda lye (36B) and recovered several times with ether, then washed in water, dried with sodium anhydrous sulfate, and then distilled. 32 G of ptrifluoromethylaniline are obtained. (Yield: about 100 percent m.p.: 32C) Stage D P-trifluoromethylphenol Into a 1 liter three-necked flask equipped with a mechanical agitator, a thermometer and a dropping funnel there are introduced 180 ml of water and then 27 ml of sulfuric acid (d: 1.84), With the temperature maintained at about 20C, 32 g (0.2 mole) of p-trifluoromethylaniline are poured in slowly, and then drop by drop, a solution of 15 g of sodium nitrite in 70 ml of water. The mixture is maintained under agitation at room temperature for about 1 hour. The product formed is steam distilled and then recovered several times in ether. The ether is dried and distilled. 27.5 G of p-, trifluoromethylphenol are obtained. (Yield: percent) Stage E P-Trifluoromethylphenoxyisobutyric acid Into a 1 liter three-necked flask equipped with a mechanical agitator, an ascendant refrigerant and a dropping funnel there are introduced 27.5 g (0.17 mole) of p-trifluoromethylphenol, 105 g of acetone and 37 g of pellets of soda. The mixture is refluxed and 25 g of chloroform are poured slowly through the dropping funnel. When the reaction is completed, the acetone is removed under vacuum and the residue recovered in water and the pH is raised to 1 with concentrated hydrochloric acid. The product obtained is extracted in chloroform. 21 G of p-trifluoromethylphenoxyisobutyric acid are obtained. (Yield: 50 percent m.p.: C)

Stage F Ethyl P-Trifluoromethylphenoxyisobutyrate Into a 500 m1 flask equipped with an asecndant refrigerant there are introduced 21 g (0.085 mole) of p-trifluoromethylphenoxyisobutyric acid and a mixture of ml of absolute ethanol and 4 g of concentrated sulfuric acid of 66B. The mixture is refluxed for 5 hours. It is extracted in chloroform, the solvent removed and the residue distilled. 13.8 G of ethyl p-trifluoromethylphenoxyisobutyrate are obtained. (Yield: 60 percent boiling at 2 to 3 mm.: 9l94C) Other examples of compounds useful in the practice of the methods of treatment of this invention are the corresponding methyl or isobutyl ptrifluoromethylphenoxyisobutyric acid. Such corresponding compounds are produced by employing instead of 100 ml of ethanol, 70 ml of anhydrous methanol or 160 ml of isobutyl alcohol in Stage F.

Each of the active products of this invention may be incorporated in a tablet, capsule, ampule or other pharmaceutical form for administration to a mammal for the treatment of cholesterolemia or biliary lithiasis. For example, the active product, such as ethyl-P- trifluoromethylphenoxyisobutyrate, may be combined with pharmaceutical carriers such as lactose, starch and a small amount of magnesium stearate when incorporated in a tablet. Any one of the other active products described in Examples 11, III and IV may likewise be combined with suitable pharmaceutical carriers in a tablet or other dosage form for administration to the mammal being treated.

EXAMPLE II ETHYL-P-TR1FLUOROMETHYLTHIOPHENOX- YISOBUTYRATE Stage A P-Nitrophenylmethylsulphide Into a liter flask equipped with a mechanical agitator, an ascendant refrigerant and a dropping funnel there are introduced 315 g. (2 moles) of pchloronitrobenzene and 1,500 ml. of 95 percent ethyl alcohol. The mixture is refluxed with agitation. As the reflux is continued, there is added in a thin stream a solution of 350 g. of sodium sulfide-Q H 0, 50 g. of sulphur and 2,000 ml. of 95 percent alcohol.

Reflux is continued for one-half hour and then a solution of soda pellets in 150 ml. of water and 500 ml. of 95 percent alcohol is poured into the reaction mixture. x i 1 The reaction product is poured intoa mixture of ice and water. The impurities are eliminated by filtration. The filtrate is acidified with hydrochloric acid (d: 1215 Theclear yellow precipitate obtained is dried and washed with water. It is then dissolved in 200 ml. of soda lye (36 B) and 2,000 ml. of water and poured into a 5 liter flask equipped with a mechanical agitator, an ascendant refrigerant and a dropping funnel. Then under agitation, 380 g. of dimethyl sulfate are added. The mixture is refluxed for 1 hour, being assured that the medium is alkaline. The mixture is then allowed to return to room temperature. The product crystallizes, is dried, washed with water and dried at 50 C. After recrystallization in methanol, 16 g. of p-nitrophenylmethrecrystallized in acetone. 218 G. of pnitrophenyltrichloromethylsulfide are obtained (Yield: percent, m.p. 94 C.) Stage C P-Nitrophenyltrifluoromethylsulphide Into a 1 liter flask equipped with a small distillation column and a descendant refrigerant, are introduced 218 g. (0.8 mole) of p-nitrophenyltrichloromethylsulphide and 218 g. of pulverized antimony trifluoride. The mixture of the two compositions is brought rapidly to the boiling point. 1t is then distilled under vacuum. The product obtained is recovered in 900 ml. of sulfuric ether and ml. of hydrochloric acid (d: l .19) diluted one-half. The etherified layer is decanted and recovered several times by washing in one-half hydrochloric acid. The ether is dried in anhydrous sodium sulphate and then distilled. The product is then distilled. 1 12.4 G. of p-nitrophenyltrifluoromethylsulflde are obtained. (Yield: 60 percent, boiling at 9 mm.: 99103 C.) Stage D P-Aminophenyltrifluoromethylsulfide Into a 6 liter three-necked flask equipped with a mechanical agitator, a thermometer and a dropping funnel, are introduced 112.4 g. (0.5 mole) of pnitrophenyltrifluoromethylsulfide and 400 ml. of 95 percent ethyl alcohol. Reduction is effected with a solution of 650 g. of stannous chloride in 490 ml. of hydrochloric acid ((1: 1.19). After neutralization in soda, the product is extracted several times in ether, which is washed in water, dried in anhydrous sodium sulfate and then distilled. The product is then distilled. 80 G. ofp-aminophenyltrifluoromethylsulphide are obtained. (Yield: 83 percent, boiling at 10 mm.: 98-99 C.) Stage E P-Hydroxyphenyltrifluoromethylsulphide Into a 2 liter three-necked'flask equpped with a mechanical agitator, a thermometer and a dropping funnel, there are introduced 450 ml. of water and then 100 ml. of pure sulfuric acid (d: 1.84). With the temperature being maintained atj20 C., 80g. of p-aminophenyltrifluoromethylsulphide are poured in, in a'th'in stream, and then, drop by drop, a solution of "40 g. of sodiumnitrite in ml. of water. The mixture is maintained under agitation at room temperature for about 1 hour. The product formed is drawn off by steam and the product is recovered several times in ether. The ether is dried and distilled. 68.5 G. of p-hydroxyphenyltrifluoromethylsulphide are obtained. (Yield: 85 percent, m.p.: 52 C., boiling at 10 mm.: 90-100 C.) Stage F P-Trifluoromethylthiophenoxyisobutyric acid Into a 1 liter three-necked flask, equipped with a mechanical agitator, a reflux refrigerant and a dropping funnel, there are introduced 260 g. of acetone, 68.5 g. of p-trifluoromethylthiophenol and 100 g. of soda pellets. The mixture is refluxed and 70 g. of chloroform are poured slowly into the dropping funnel. When the reaction is complete, the acetone is removed under vacuum and the residue recovered in water, then brought to a pH of 1 with concentrated hydrochloric acid. The product obtained is extracted with chloroform. 69 G. of ptrifluoromethylthiophenoxyisobutyric acid are obtained. (Yield: 70 percent, m.p.: 118 C.) Stage G Ethyl P-Trifluoromethylthiophenoxyisobutyrate Into a 500 ml. flask equipped with an ascendant refrigerant, are introduced 69 g. of ptrifluoromethylthiophenoxyisobutyric acid and a mixture of 280 ml. of absolute ethyl alcohol and 12 g. of sulfuric acid of 66 B. The mixture was re fluxed for 6 hours. The mixture was extracted with chloroform, the solvent removed and the residue distilled. 49 G. of ethyl p-trifluoromethylthiophenoxyisobutyrate are obtained. (Yield: 65 percent, boiling at 5 mm.: 1145 C.)

Other compounds useful in the practice of the methods of treatment of this invention are the corresponding isopropyl or n-pentyl p-trifluoromethylthiophenoxyisobutyrate. Such corresponding compounds are produced by employing instead of 100 ml of ethanol, 130 ml of anhydrous isopropyl or 190 ml of l-pentyl alcohol.

EXAMPLE 111 ETHYL P-TRlFLUOROMETHYLSULFINYLPHENOX- YISOBUTYRATE Stages A through F are the same as those described in Example 11.

Stage G P-Trifluoromethylsulfinylphenoxyisobutyric acid into a 2 liter three-necked flask, equipped with a mechanical agitator, a thermometer and a dropping funnel, are introduced 800 ml. of methanol, a solution of 40 g. of sodium metaperiodate in 370 ml. of water and finally 50 g. (0.178 mole) of ptrifluoromethylthiophenoxyisobutyric acid. The mixture is maintained under agitation at room temperature for 4 /2 hours. When the reaction is completed, the product, having been recovered several times in chloroform, is extracted. 44 G. of ptrifluoromethylsulfinylphenoxyisobutyric acid are obtained. (Yield: 83 percent, m.p. 123 C.)

Stage H Ethyl P Trifluoromethylsulfinylphenoxyisobutyrate The esterification of p-trifluoromethylsulfinylphenoxyisobutyric acid is effected as described in Example 11, stage G. The corresponding ester is obtained. (Yield: 58 percent, boiling at 5 mm.: 122l24 C.)

EXAMPLE 1V.

ETH YL P TRIFLUOROMETHYLSULFONYLPHENOX- YISOBUTYRATE Stages A through F are the same as those described in Example 11. Stage G P-Trifluoromethylsulfonylphenoxyisobutyric Into a 1 liter three-necked flask equipped with a mechanical agitator and an ascendant refrigerant, are introduced 58 g. of p-trifluoromethylthiophenoxyisobutyric acid dissolved in 300 ml. of acetic acid and 125 ml. of hydrogen peroxide in l 10 volumes. When the reaction is completed, the acetic acid is dissolved under vacuum. The crystallized product at the bottom of the flask is recovered by water, dried, washed several times with water, then dried at 50 C. G. ofp-trifluoromethylsulfonylphenoxyisobutyric acid are obtained. (Yield: percent, m.p.: 101 C.)

Stage H Ethyl P-Trifluoromethylsulfonylphenoxyisobutyrate The esterification of p-trifluoromethylsulfonylphenoxyisobutyric acid is effected as illustrated in Example 11, stage G. Ethyl ptrifluoromethylsulfonylphenoxyisobutyrate is obtained. (Yield: 61 percent, boiling at 5 mm.: 149152 C.) The compositions of this invention have been the subject of pharmacological and clinical studies to determine both their non-toxicity, their activity in controlling cholesterolemia and in preventing biliary lithiasis.

The toxicity tests of the medicaments of this invention carried out on mice weighing an average of 22 g. showed perfect tolerance of doses up to mg. per mouse. (one-tenth ml.)

The pharmocological studies of the medicaments of the invention showed significant protection against lithiasis of the gall bladder as well as protection against chlesterolemia.

The preventive action in lithiasis of the gall bladder was studied in accordance with the test of Francois Besancon and C011. described in Socie't Mdicale des Hopitaux de Paris 1,966, 117, no. 2, 127-138.

Four lots of five female mice 3 months old, of Swiss- Gif breed were tested by lithogenous regime for 6 weeks. The lithogenous regime is a normal nourishment for mice supplied by the U.A.R. Establishments, with the addition of 0.5 percent dosage of sodium dehydrochlorate (cf. Francois Besancon et a1. 1965-1966).

The lot tested received no other treatment.

One of the medicaments of the invention was administered to each of 3 other lots subcutaneously in dosages of mg 6 days per week for 6 weeks.

At the end of the treatment, the mice were sacrificed and autopsies performed on them. The weight'of the animals, the weight and color of their livers, the dimensions of the gall bladder, the presence of macroscopically visible stones and observations by polarizing microscope were noted. The findings are given in the fol- 5 acid 5 lowing table.

TABLE 1 TREAT- WElGH'lWElGHT PERCENT COLOR OF VESlCLE BlREFRlNGENT MENT OF OF WEIGHT LIVER LlTHl- DlMEN- CRYSTALS LlVER OF ASIS SlONS OUSE IN G. IN G. LIVER mm.

None 30 1.68 5.6 colorless 7X3 numerous large masses None 28 1.67 6 normal 5.5x normal 2.5 None 28 1.38 4.9 normal 6X3 many large masses Table l-Continued TREA'F WEIGH'IWEIGHT PERCENT COLOR OF VESICLE BIREFRINGENT MENT OF OF WEIGHT LIVER LITHI- DIMEN- CRYSTALS LIVER OF ASIS SIONS OUSE IN O. IN (E. LIVER mm.

None 35 2 5.7 normal 6.5x normal 3 None 25 1.04 4.2 normal 7.5X many large masses 3 normal Compound 30 3.25 11 normal 6X2.5 normal 20 2.60 13 normal 0 4X1 normal 30 2.30 7.7 normal 0 6X3 normal Compound 22 3.85 18 normal 0 6X2 small no. of II large masses many small masses 20 3.50 [8 normal 0 11X2 normal 24 3.60 15 normal 0 5X1 .5 numerous very small masses Compound 25 4.80 19 normal 0 6X25 numerous very 111 small masses 21 3.92 19 color- 0 7.5 4 normal less 3.70 19 normal 0 7X2 normal Compound I ethyl p-trifluoromethylsulfonylphennxyisnbulyrate Compound llethyl p-lril'luoromclhylthiophenoxyisohutyrate Compound lll-ethyl p-tril'lunromclhylphcnoxyisohutyratc Table I demonstrates that in the five test mice subjected to the lithogenous regime with no other treatment, four showed macroscopically visible lithiasis and the polarizing microscope showed large cohesive doubly refractive crystals. In the fifth mouse, microscopic study showed normal bile and absence of lithiasis. The normal aspect of the bile is demonstrated by the existence of few doubly refractive crystals of small size and a maximum of 2 or 3 larger crystals in the gall bladder content.

The animals (male rats WISTAR) received orally a combination of androsterone and the medication unde r study incorporated in the diet in rates of 0.01 percent and 0.25 percent respectively for 11 days.

At the conclusion of the experiment, the rats were vention are given in the following table.

TABLE II" TREAT- NO. OF 3 AVERAGE WEIGHT IN G. GAIN DAILY CHOLES- MENT ANIMALS BEFORE AFTER TESTING INTAKE TEROLEM- TESTING OF IA G/L NOURISH- MENT IN '0 5 162 226 39% f 81 0.92 Androsterone 5 187 249 33% 97 0.90 Androsterone 4 I86 270 89 0.76 Compound I Androslterone 5 151 198 31% 85 0.63 Compound 11 Androsterone 5 168 208 23% 83 0.61

Compound III "One animal losing weight because of a microbial infection was eliminated.

On the other hand, in the animals treated by the Ii- The comparison of the cholesterolemias of the anithogenous regime supplemented by the medications of this invention, no lithiasis was observed. Only 2 mice showed small crystals under microscopic examination, but in greater than normal numbers.

The other mice were normal under microscopic examination.

Thus the significant efficacy of the medicaments of this invention against gall bladder lithiasis is shown experimentally.

The hypocholesterol control activity of the compositions of this invention were studied according to the technique of THORP and WARING (Nature 1,962, 194, 948-49) as described hereafter.

mals treated with androsterone or in combination with compounds of the invention was made by comparative analysis. The results show a highlysignificant hydrocholesterolemia action by three of the compounds of this invention.

Tests were conducted to determine the lethal dosage in mice of compounds used in the methods of treatment of this invention. The compounds tested were:

A ethyl p-trifluoromethylthiophenoxyisobutyrate The lethal dose is therefore above 320 mg/kg.

The compounds of this invention therefore are entirely safe to use in daily dosage of to 360 mg/kg of body weight.

The pharmacological testing shows that the methods of treatment of this invention are safe and effective in the treatment of cholesterolemia and suppress biliary lithiasis in mammals.

The pharmacological results were confined in human therapy, the compositions being administered in the form of capsules in dosages ranging from 1 to 2.5 grams per day.

What is claimed is:

COOR

C Ila-(f-Clh in which R is a straight or branched chain alkyl having 1 to 5 carbon atoms and X is trifluoromethyl, trifluoromethylthio, trifluoromethylsulfinyl or trifluoromethylsulfonyl.

2. The method of claim 1 in which the daily dosage is to 280 mg/kg of body weight.

3. The method of claim 1 in which the fluorinated derivative is ethyl p-trifluoromethylphenoxyisobutyrate.

4. The method of claim 1 in which the fluorinated derivative is ethyl p-trifluoromethylthiophenoxyisobutyrate.

5. The method of claim 1 in which the fluorinated derivative is ethyl p-trifluoromethylsulfinylphenoxyixobutyrate.

6. The method of claim 1 in which the fluorinated derivative is ethyl p-trifluoromethylsulfonylphenoxyisobutyrate. 

2. The method of claim 1 in which the daily dosage is 140 to 280 mg/kg of body weight.
 3. The method of claim 1 in which the fluorinated derivative is ethyl p-trifluoromethylphenoxyisobutyrate.
 4. The method of claim 1 in which the fluorinated derivative is ethyl p-trifluoromethylthiophenoxyisobutyrate.
 5. The method of claim 1 in which the fluorinated derivative is ethyl p-trifluoromethylsulfinylphenoxyixobutyrate.
 6. The method of claim 1 in which the fluorinated derivative is ethyl p-trifluoromethylsulfonylphenoxyisobutyrate. 